Ionization Properties of Phospholipids Determined by Zeta Potential Measurements
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Author:
Date:
2016-11-20
[Abstract] Biological membranes are vital for diverse cellular functions such as maintaining cell and organelle structure, selective permeability, active transport, and signaling. The surface charge of the membrane bilayer plays a critical role in these myriad processes. For most biomembranes, the surface charge of anionic phospholipids contributes to the negative surface charge density within the interfacial region of the bilayer. To quantify surface charge, it is essential to understand the proton dissociation behavior of the titratable headgroups within such lipids. We describe a protocol that uses ...
[摘要] 生物膜对于多种细胞功能如维持细胞和细胞器结构,选择性渗透性,主动转运和信号传导至关重要。膜双层的表面电荷在这些无数过程中起关键作用。对于大多数生物膜,阴离子磷脂的表面电荷有助于在双层的界面区域内的负表面电荷密度。为了量化表面电荷,必须理解可滴定头基在这种脂质内的质子解离行为。我们描述了使用模型膜用于电动ζ电位测量以及使用Gouy-Chapman-Stern形式的数据分析以确定组分脂质的p a 值的方案。提供了由单阴离子脂质磷脂酰甘油组成的均匀双层的详细实施例。这种方法可以适用于测量具有异质脂质组合的双层,以及用于在头部组中具有多个可滴定位点的脂质(例如,心磷脂)。 [背景] 磷脂是生物膜的中心结构单元(图1)。作为两亲性分子,每个包含由酰基链组成的疏水区和由极性头基组成的亲水区(图1A)。一些磷脂头基是两性离子的,在生理pH下含有带正电和带负电的官能团(图1B),而其它磷脂头基是酸性的,带有整体形式的负电荷(图1C)。生物膜内的脂质作为层状组件稳定存在,形成双层,其中两个叶的酰基链相互作用形成疏水核和由极性头基组成的两个界面区域(图1D)。大多数天然存在的生物膜含有一定百分比的酸性磷脂;因此,它们的脂质组成赋予界面区域净负电荷(Gennis,1989; ...
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Quantitative Analysis of Cellular Diacylglycerol Content
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Author:
Date:
2014-08-05
[Abstract] Diacylglycerol (DAG) is a bioactive lipid with diverse biological roles. DAG transiently accumulates in a membrane upon receipt of an appropriate stimulus that activates phospholipase C to cleave phospholipids. The resulting hydrolysis product DAG binds to proteins such as protein kinase C to initiate a variety of downstream cellular processes. DAG kinases attenuate such responses by converting DAG to phosphatidic acid.
This protocol describes an assay designed to quantify cellular DAG levels. The assay exploits the enzymatic conversion of DAG (sn-1,2-diacylglycerol) to ...
[摘要] 二酰基甘油(DAG)是具有不同生物学作用的生物活性脂质。 DAG在接受激活磷脂酶C以切割磷脂的适当刺激时在膜中瞬时累积。 所得水解产物DAG结合蛋白质例如蛋白激酶C以启动各种下游细胞过程。 DAG激酶通过将DAG转化为磷脂酸来减弱这种应答。该方案描述了设计用于定量细胞DAG水平的测定。 该测定利用DAG(1,2-二酰基甘油)到磷脂酸(1,2-二酰基-sn-甘油-3-磷酸)的酶促转化 结合DAG激酶掺入放射性标记的磷酸基团(图1)。 该测定在(Strijbis等人,2013)中描述。
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